RNA lifetime and abundance is regulated by shifting the balance between RNA transcription and RNA degradation. Two principle RNA decay pathways exist in eukaryotes, one catalyzes degradation 5' to 3' while the other degrades RNA in the 3' to 5' direction. The 3' to 5' decay pathway requires the activities of the RNA exosome, a large multi-subunit protein complex that contains a non-catalytic core of nine subunits and two additional subunits that catalyze processive and distributive 3' to 5' RNA exoribonuclease activities. In budding yeast, ten of the eleven genes are essential for growth, suggesting the importance of the RNA exosome and its activities in cellular function. While recent efforts illuminated fundamental aspects of eukaryotic exosome structure and function, many questions remain with respect to the individual and collective activities for exosome subunits in RNA processing and decay. RNA decay pathways play an integral role in eukaryotic nucleic acid metabolism, so our studies are of direct relevance to human health and the mission of the NIH because misregulation of RNA processing and decay is associated with disease states such as cancer, inflammation and neurodegenerative diseases. This proposal will address the central issues of human and yeast RNA exosome biology by characterizing individual exosome subunits, by reconstituting multi-subunit RNA exosomes and by analyzing the activities of these complexes in biochemical, genetic and structural studies that will establish functions for the RNA exosome during RNA degradation in vitro and in vivo. PUBLIC HEALTH RELEVANCE: The eukaryotic RNA exosome is an essential complex that regulates RNA homeostasis through its 3' to 5' processing and decay activities. RNA exosomes also contribute to RNA surveillance via quality control pathways that target aberrant RNA molecules for destruction. Together, these pathways regulate the lifetime of particular RNAs and thus serve a vital role in signal transduction by controlling the duration of transient bursts of RNA expression that are associated with signaling cascades. The RNA exosome also protects the cell from deleterious RNA that could lead to cellular pathology. Defects in these pathways are associated with several human diseases including cancer, inflammation, and neurodegenerative disorders.